Cultivation of animal cells, especially in large quantities, is essential to the production of antiviral agents such as viral vaccines and interferon or biologically active chemicals such as hormones. In particular, the production of monoclonal antibodies having the ability to bind to a specified protein as a target relies on the cultivation of a large quantity of a hybridoma obtained by fusing antibody-producing cells with myeloma cells, and the solution of problems associated with this technique is an industrially important subject.
Heretofore, cell culture has been carried out on a laboratory scale by using a Petri dish, a test tube, a cultivation bottle, etc. Generally, the cell culture is classified into anchorage dependent cell culture and suspension culture depending upon the cells to be cultivated. The suspension culture would be an industrially advantageous method since it has a possibility of cultivating cells at a high density.
Cell cultures usually demand supply of oxygen (O.sub.2), and for this purpose, the concentration of oxygen in the suspension is maintained constant by, for example, supplying an oxygen-containing gas from the gaseous phase above the liquid surface of the suspension and dissolving it in the suspension, or by blowing an oxygen-containing gas into the suspension. The supply of oxygen by these methods does not particularly give rise to any problem in the cultivation of cells on a small scale.
But where cell culture is desired to be effected on an industrial scale, above all at a high cell density, all of the above methods of oxygen supply are unsuitable. In the case of supplying oxygen from the free liquid surface of the suspension, even when the amount of the suspension increases, the area of the liquid surface cannot be increased correspondingly. Thus, on an industrial scale, it is almost impossible to avoid the insufficiency of oxygen supply.
When the oxygen-containing gas is blown into the suspension, the liquid surface rises owing to bubbling, and it is sometimes even difficult to continue the operation. Furthermore, this method is difficult to apply to cells which might die or decrease in proliferating activity upon contact with bubbles, or to cells which undergo a separation phenomenon by the action of bubbles (a certain kind of plant cells, for example).
Recently, a different method was proposed in Japanese Patent Publication No. 4235/1982. This patent document discloses a method of cultivating cells which comprises adhering cells to the surface of a semipermeable containing a quaternary ammonium salt having a water content of 20 to 90%, and passing a cultivation fluid over at least the opposite side to the cell-adhering surface of the membrane. This method is characterized by the fact that nutrients and oxygen are supplied to the cells through the semipermeable membrane without direct contact of the cells with the cultivation fluid.
The cultivation of plant cells has the same problem as the cultivation of animal cells. It is known that the cultivation of plant cells can give the same substances as primary or secondary metabolites obtained from the parent plants, such as enzymes, terpenoids, flavonoids, steroids, alkaloids, quinones and phenols, and useful substances in the fields of medicines, foods, cosmetics and fine chemicals. It is very important therefore to solve the same problem of the cultivation of plant cells as in the case of the cultivation of animal cells.
Science, volume 219, pages 1448-1449, March 1983 discloses a method in which a fluorocarbon emulsified and stabilized with polylysine is used as a microcarrier, and anchorage-dependent cells are cultivated on the microcarrier.
U. S. Pat. No. 3,850,753 discloses a method of cultivating an aerobic microorganism in the presence of a water-immisicible inert liquid fluorocarbon under aeration, agitation and/or shaking.
It is an object of this invention to provide a novel method of suspension cultivation on an industrial scale of animal and plant cells.